Abstract

Human ACTH and a number of ACTH peptide fragments were prepared by solid-phase synthesis. Of the analogues tested, only ACTH_1-39 and ACTH_1-24 stimulated rat adrenal membrane adenylate cyclase in vitro. Several shorter fragments, including ACTH_6-39, ACTH_9-24, ACTH_9-19 amide, ACTH_9-20, ACTH_9-18, and ACTH_12-39, inhibited stimulation of the enzyme by native bovine ACTH. The inhibition produced by this group of peptides appears to be specific for ACTH-stimulated adrenal adenylate cyclase. Basal and fluoride-stimulated enzyme activity in adrenal and fluoride- or epinephrine-stimulated activity in liver were not inhibited by ACTH_9-24. To elucidate further the action of the inhibitory peptides, we tested their ability to inhibit the binding of [¹²⁵I]ACTH_1-24 to adrenal particles. Peptides having either agonist or antagonist activity in the adenylate cyclase assay were effective competitors for binding. The order of potency according to both adenylate cyclase inhibition and competitive binding affinity was ACTH_6-39 > ACTH_9-19 amide ≅ ACTH_9-24 > ACTH_9-20 ≅ ACTH_9-18 > ACTH_12-39. The analogues ACTH_1-8, ACTH_9-16, and ACTH_19-39 were inactive in both the adenylate cyclase and binding systems. Comparison of this series of closely related peptides in two independent assay systems should more clearly define the structural requirements for the binding of ACTH to its membrane receptor.